Effect of different retting processes on yield and ...

WJNF #1401505, VOL 00, ISS 00

Effect of different retting processes on yield and quality of banana pseudostem fiber R Brindha, C.K Narayana, V Vijayalakshmi, and R.P Nachane

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JOURNAL OF NATURAL FIBERS 2017, VOL. 00, NO. 00, 1–10 https://doi.org/10.1080/15440478.2017.1401505

Effect of different retting processes on yield and quality of banana pseudostem fiber R Brindhaa, C.K Narayanab, V Vijayalakshmia, and R.P Nachanec a

Department of Biotechnology, Bharathidasan Institute of Technology, Anna University, Tiruchirappalli, India; Division of PHT, ICAR-Indian Institute of Horticultural Research, Bengaluru, India; cDivision of Quality Evaluation and Improvement, ICAR-Central Institute for Research on Cotton Technology, Mumbai, India

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ABSTRACT

KEYWORDS

The main objective of this study is to develop a suitable technology to utilize banana pseudostem waste in an effective manner. The choice of a specific extraction method depends on the intended end uses of the fibers and hence different methods (mechanical, microbial, chemical, and enzymatic) were carried out to extract cellulosic fibers from Poovan variety and the chemical properties were investigated. The flexural and tensile properties of fibers were explored to analyze the suitability of fibers for different applications. Results obtained from these analyses confirmed that the tex value of chemical retted fiber was lesser than others. Scanning electron microscope micrographs revealed that the surfaces of the chemically retted fibers were rougher than mechanically extracted fibers with an average diameter of 180 µm. The elemental composition of the chemical-treated banana pseudostem fibers was investigated by energy-dispersive X-ray analysis. The Fourier transform infrared spectroscopy spectrum indicated the presence of similar functional groups in all the fiber samples.

Banana pseudostem; fiber quality; microstructure; retting methods; tenacity; Fourier transform infrared spectroscopy (FTIR)

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关键词

香蕉假茎; 纤维品质; 微结 构; 脱胶方法; 韧性; 傅立 叶变换红外光谱 (FTIR)。

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摘要

本研究的主要目的是开发一种有效利用香蕉伪茎废料的技术。一个具体 的提取方法的选择取决于最终的用途，因此纤维不同的方法（机械、生 物、化学和酶法）进行了从Poovan品种和化学性质进行了提取的纤维素 纤维。探讨了纤维的弯曲和拉伸性能，分析了纤维在不同应用场合的适 用性。从这些分析得到的结果证实，化学浸渍纤维特克斯值较小。SEM 照片显示化学沤制的纤维表面比粗糙机械提取纤维的180µM.能量色散X 射线分析研究了处理香蕉假茎纤维的化学元素组成的平均直径。FTIR光 谱表明在所有的纤维样品中都存在类似的官能团。

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Introduction Plant fibers are lignocellulosic materials and have prospective industrial applications in polymer composites, textile, and paper industries. The banana plant has been one of the oldest sources of natural fiber exploited by mankind for tying flower garlands and for making crafts like baskets, bags, and more. Banana pseudostem fiber (BPF) is a bast fiber that can be extracted after the harvest of fruit bunch, and its fiber content is approximately 54.3% fibers (Ganan et al. 2004). The specific characteristics of BPFs such as high absorbance, good thermal stability, and biodegradability make them more useful in industries than other natural fibers. Banana fibers are extensively used as raw material for paper cardboards, currency notes, high-quality textiles, and reinforced as polymer

CONTACT R Brindha [email protected]; [email protected] Department of Biotechnology, Bharathidasan Institute of Technology, Tiruchirappalli 620 024, India Color versions of one or more of the figures in the article can be found online at www.tandfonline.com/WJNF. © 2017 Taylor & Francis

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composites (Chauhan and Sharma 2014; Reddy and Yang 2005; Salit 2014) and as major ingredient in Green-Compressed Earth Block (Mostafa and Uddin 2015). In spite of its wide applications, this fiber biomass is usually dumped in the soil of the banana-growing areas for enrichment, which encourages the breeding of banana pests and causes additional environmental problems. Hence, its economic utilization can bring benefits to the farming community. The separation of banana fiber from its biological matrix, with retention of quality in order to enable its suitability for different uses, is the major challenge even though physical (hand stripping) and mechanical (hammering, decortications) methods are already well explored. The deterioration of mechanical property, incomplete removal of vegetable matter, and low yield of the fiber output are the serious limitations in these methods. Banana fiber can also be extracted by retting, as in case of other natural fibers like jute and hemp. However, in the conventional retting process, the quality of fiber is reported to be adversely affected by under-retting or over-retting (Ganan et al. 2004), and therefore requires expert supervision. Very little research work has been carried out in evaluating the properties of banana fiber, which would help in its value-added applications. To the best of our knowledge, this is the first report assessing four different retting processes from the point of view of its industrial application. Therefore, the present study was undertaken to carry out different processes for the extraction of BPFs, and its effect on fiber yield, surface morphology, physical, chemical, and mechanical properties.

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Materials and methods Materials The pseudostem of banana (cultivar Poovan) was obtained from the ICAR-National Research Centre for Banana, Tiruchirapalli. Sodium hydroxide, ethylene diamine tetraacetic acid (EDTA), sodium phosphate, and oxalic acid were purchased from Himedia and commercial grade enzyme Pectinase plus was purchased from Biocon India limited, Bangalore, India. Aspergillus niger was obtained from the National Collection of Industrial Microorganisms, Pune, India, and was maintained on potato dextrose agar medium slants for 4° C. For long-term storage, glycerol stocks were prepared and were preserved at −80°C. Extraction methods Defect-free fresh banana pseudostem sheaths with dimensions of approximately 30.5 cm length, 8 cm width, and 1cm thickness were used for experimentation. About 3 kg of these fresh sheaths were soaked in tanks with 20 L of distilled water whilst ensuring their complete immersion. A. niger inoculum was prepared by transferring a loop full of culture from the agar slant into a 250 mL Erlenmeyer flask containing 100 mL potato dextrose broth incubated in an orbatory shaker at 200 rpm at 30°C at pH 4 until the phase of spore formation. About 1.25% of fungal spore solution (1 mL contained 1.28 × 105 spores) was inoculated into the tank to which 0.1% ammonium dihydrogen phosphate and 0.5% glucose were supplemented for growth enhancement. The set-up was kept undisturbed for several days with intermediate examination for complete retting. The control sample consisted of pseudostem sheaths of same dimension in tanks with groundwater obtained from the ICAR-NRCB farm. The water used contains innate microorganisms, and it was not further enriched with additional carbon and nitrogen sources. For enzymatic retting treatment, a tank containing distilled water with 0.2% (v/v) commercial pectinase was used, in which the pseudostem sheaths were immersed. The pH was adjusted to 5.0– 5.5 in order to maintain maximum enzyme activity and the setup were left at ambient temperature (28–40°C). Samples were checked intermittently for completion of the retting process. For chemical retting, pseudostems were treated with sodium hydroxide (0.5%, 1%, 2%); oxalic acid (0.5%, 1%, 2%), and a combination of EDTA, sodium phosphate, and NaOH at 8, 50, and 25 mM, respectively, for 4 h. Two different conditions were used: (i) immersion at room temperature 37°C and (ii) immersion in water bath at 97°C. Physical observations were made periodically to confirm the

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completion of retting process. After retting process, each fiber sample was cleaned with distilled water to reduce the presence of vegetable residues and dried in hot air oven at 80°C for 2 h prior to analysis.

Yield and chemical analysis Except in the chemical retting process, the pseudostem samples under treatment were withdrawn at a regular interval of 6 days and the changes in cellulose, starch, lignin, and pectin were evaluated. After extraction process, the fiber yield was calculated using Eq. (1). Yield% ¼

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Wf  100 Ws

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where Wf is the weight of fiber obtained after retting process, and Ws is the weight of pseudostem sheath initially taken. Cellulose, an important component of fiber and pseudostem, was estimated using the method suggested by Updegraff (1969). Starch were estimated using the method of Hodge and Hofreiter (1962). Acid insoluble lignin content was measured in accordance with TAPPI standard T222 om-02 (Effland 1977). Pectin was estimated as calcium pectate adopting a procedure outlined by Ranganna (1986) .

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Fiber characterization The structural composition and nature of the functional groups of the fiber samples obtained after 100 extraction process were studied by Fourier transform infrared spectroscopy (FTIR) analysis. FTIR spectra were recorded on Perkin Elmer FTIR spectrophotometer-spectrum RX1 with a scan rate of 20 scans per minute and a resolution of 4 cm−1 in the wavelength region from 4000 to 400 cm−1. A field emission scanning electron microscope (FE-SEM) (ZEISS SIGMA VP) was employed to study the surface morphology of fiber samples. Energy-dispersive X-ray (EDX) analysis was also 105 carried out using Xflash 5030 Silicon Drift Detector to study the chemical composition and the distribution of the chemical elements of interest in fiber samples. Tensile and flexural properties of fiber samples were analyzed in Quality Evaluation and Improvement Division at ICAR-Central Institute for Research on Cotton Technology, Mumbai. Properties of fibers such as load at max, % strain at max, Young’s modulus, energy to break point, 110 tex, and tenacity were analyzed using standard procedures.

Results and discussion Anatomical characteristics of banana pseudostem Anatomical characteristics of banana pseudostem were determined in order to observe the position of fiber bundles in the matrix. Free hand transverse sections of banana pseudostem were made and 115 viewed under optical microscope (ECLIPSE TS100, Nikon, Japan) at 10× and 40× magnifications. Fiber bundles were seen to be distributed throughout both in surface and middle lamella regions. The vascular bundles of BPF were widely distributed in parenchymatous ground tissue, which consisted only of fiber and phloem, without any other vascular tissues (Figure 1). This situation might occur in certain types of monocot stem. In plant tissue, phloem plays an important role in 120 organic nutrient transport, especially in the case of sugar that is produced during the photosynthesis process. Similar results were obtained in the analysis of morphological structure of banana pseudostem by Li et al. (2010).

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Figure 1. Free hand transverse section of banana pseudostem after stained with lacto phenol blue at 10× and 40× magnification. P, parenchyma cell; Ph, phloem cell; F, fiber cell.

Extraction of fiber Fiber extraction from banana pseudostem was carried out by various methods such as mechanical extraction (using a decorticator), chemical, microbial, and enzymatic retting processes. Among the retting processes studied, the treatment using innate microorganisms in groundwater showed good retting at the end of 18th day, whereas treatment using A. niger took 22 days to show good results. The presence of mixed microbial populations in groundwater could ferment the sheath and release the cellulosic fibers at an accelerated rate when compared with fungal retting even though the retting medium of fungus was enriched with glucose and ammonium dihydrogen phosphate. In the absence of additional carbon and nitrogen sources, innate microbes utilized the pseudostem biomass as substrates for its growth, which would have accelerated the retting process. These results confirmed that the retting period is independent of retting medium composition, amount of inoculum added, and the type of microbial strain used for retting. Similar results were well documented by Maheshwari et al. (1994) and Sarma and Deka (2016). Pectin plays a major role in holding the fibers together in the outer walls of banana fiber bundles (Ganan et al. 2004). Since the separation of fiber bundles is easily accomplished by the removal of pectin from the sheath, enzyme pectinase, and A. niger, a known pectinase producer was also employed in the retting methods. Both the treatments expressed low retting capacity due to the specific degradation of pectin in the pseudostems. Among various trials of chemical retting, boiling the sheath with NaOH at the concentration of 0.5% and 1% for 55 and 30 min, respectively, showed excellent fiber separation. Fiber samples retted by 1% of NaOH are represented herein by a general term “chemical retted” in the rest of the paper. Alkaline pH condition remarkably dissolves noncellulosic matter in jute, resulting in better fiber quality (Sikdar, Mukhopadhyay, and Mitra 1993) and hence to improvise the fiber quality, pretreatment with dilute alkali was encouraged by Padmavathi and Venkata Naidu (1998) in sisal fibers. Similarly, acidic pH of banana pseudostem retards the removal of noncellulosic material during the microbial retting process, whereas direct treatment with NaOH effectively removed plant residues and remarkably reduces the retting time from days to hours, yet chemical requirement and disposal of unused chemical in the effluent should be considered while commercially practiced. An attempt has been made by Jose et al. (2016) to reuse the retting bath by adding required quantity of chemicals to economize the chemical retting process as well as to minimize the environmental issues by virtue of effluent discharge. This strategy further aids in scaling up the process to increase the substantial yield for textile processing.

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Biochemical changes occurring during various retting processes Various biochemical parameters were determined from 0th day to 18th day to analyze the changes occurring during the microbial and enzymatic retting process. An increase in the cellulose content 155

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Figure 2. Rate of progressive change in (a) cellulose, (b) starch, (c) lignin, and (d) pectin content during retting process by different methods. Error bar represents standard deviation (n = 3). Significantly different from untreated control *P < 0.05; **P < 0.01.

during the retting process indicated the removal of noncellulosic substances from the sheath and also confirmed that cellulose was not degraded during retting process (Figure 2a). Since starch is one of the major components accumulated in the pseudostem, its removal from fiber bundles increases the efficient separation of fibers. The pseudostem had a starch content of 5.7% initially, which significantly reduced (P < 0.01) during the retting process due to its consumption by the microbes for its 160 proliferation and enzymatic action. Microbial retting showed greater decrease in starch compared to enzymatic method, indicating that the microbes released more amylase than the pectinase (Figure 2b). The lignin, though decreased slightly, did not show significant degradation during the process of retting in any of the three methods (Figure 2c). However, among the three methods natural retting using groundwater with its innate microbes showed relatively better removal of lignin 165 (throughout the treatment process P < 0.05). This indicated the presence of lignin-degrading microbes in the groundwater. In all the retting processes, pectin content decreased significantly (P < 0.01), which confirmed the degradation of pectin during the retting process (Figure 2d). Enzymatic method showed maximum decrease of pectin (61.19%) compared to other two microbial methods. From these observations, we confirm that biological retting processes progressively remove 170 the components pectin and starch, which are present in the outer region of fiber bundles but do not alter the inner elementary components especially cellulose and lignin.

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Table 1. Fiber yield and chemical composition of BPF obtained by different methods of extraction. Treatment Mechanical Innate microbes Aspergillus niger Enzyme Chemical

Fibre yield (g/kg) 5.1 ± 0.3 12.7 ± 1.2 9.5 ± 0.8 10.9 ± 0.09 9.2 ± 0.5

% Cellulose 71.33 ± 1.56 63.5 ± 2.43 62 ± 5.6 70.5 ± 2.4 65 ± 3.3

% Pectin 2.35 ± 0.02 1.775 ± 0.09 2.075 ± 0.1 1.925 ± 0.5 1.705 ± 0.1

% Lignin 17.65 ± 4.1 21.225 ± 0.94 21.9 ± 1.39 18.575 ± 1.45 12.54 ± 0.56

Yield and chemical analysis of fiber

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Table 1 depicts the yield and the chemical composition of BPF extracted from various methods. Retting by innate microbes showed the maximum fiber yield since both surface and inner elementary fibers were easily extracted due to the degradation of noncellulosic components except lignin. Chemical analysis confirmed the higher lignin content in fibers extracted by microbial retting process, which supports its applicability as reinforcement in various composites. The use of banana fiber for reinforcement shows better efficiency than coir and also its specific strength properties are equivalent to those of glass fiber-reinforced plastics (Benitez et al. 2013). It is very important to explore banana fibers in the field of fiber-reinforced composites as these are cost effective and ecofriendly than synthetic fibers like glass, carbon, and asbestos. Enzyme retted fibers showed highest cellulose content, which confirmed its applicability in paper industry (Li et al. 2010) as a good replacement for wood pulp, thereby reducing the environmental impact of deforestation. The retting method using A. niger provided comparatively lesser yield and cellulose content than other retting methods yet higher lignin content affirmed its utilization as insulators. Chemical retting method showed a twofold increase in yield without significantly affecting the strength compared with that of mechanical extraction. NaOH causes dissolution of lignin in pseudostem by breaking it into smaller segments, thereby decreasing the lignin content to a greater extent. Similar alkaline treatment is employed in paper and pulp industries for lignin removal.

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Fiber characterization Tensile and flexural properties The mechanical properties of fibers are measured in terms of their resistance to deformation under applied loads or stresses. Table 2 explicits the behavior of the fibers under gradually increasing tensile load and the parameters were evaluated from the load–elongation (stress–strain) curve. To 195 compare the fiber samples in terms of their elongation properties, % strain at max was determined. From the results (Table 2), it was confirmed that no significant difference was observed between the fibers obtained from different retting methods. To compare the fibers with different linear densities, specific strength of the fiber commonly known as tenacity was calculated from the breaking force and the linear density of the unstrained fiber specimen. Breaking force is the maximum force 200 required to break the fiber and is usually expressed in gram-force (gf). The results obtained (Table 2) confirmed that the tenacity of chemical retted fiber was quite high (75 g/tex), suggesting Table 2. Flexural properties of BPF obtained by different methods. Load at max Sample Mechanical Aspergillus niger Innate microbes Enzyme Chemical

(g) 671 651 711 548 480

CV% 28.233 27.170 34.633 34.606 36.981

Strain at max % 2.973 2.719 2.648 2.758 2.758

CV% 16.828 14.564 11.147 13.705 19.515

Slope (Aut Young) kgf/mm 0.510 0.554 0.595 0.439 0.400

CV% 20.072 26.526 32.778 34.568 32.948

Energy to break point kgfmm 0.509 0.421 0.465 0.380 0.320

CV% 40.035 34.242 37.507 37.209 45.820

Tenacity Tex 10.3 12.5 12.6 9.73 6.4

g/tex 65.02 52.08 56.43 56.32 75

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that they can replace synthetic fibers to provide better quality yarn. Low tex value (6.4) confirmed the fineness of the chemical retted sample. Initial resistance to deformation (Young’s modulus) is measured from the slope of the stress–strain curve. High slope value (0.595) indicated the stiffness 205 and low initial elongation of the fiber retted by innate microbes and hence it would be more suitable in manufacturing nonwovens. Comparable observations have been reported by Ganan et al. (2004) for banana fibers when they were treated with a consortium Fusarium spp., Trichoderma spp., acidogenic and metanogenic microbes. Significant diminution of tensile properties of microbially retted fiber bundles was observed after increasing the retting time attributing to the attack of the 210 microorganisms on the cellular walls of banana bundles (Doraiswamy and Chellamani 1993; Ganan et al. 2004). Shivashankar, Nachane, and Kalpana (2016) observed that the anaerobically retted fiber was superior over the enzymatically retted fibers in view of the tenacity of fiber. Energy required to break the fiber was high (0.509 kgf-mm) for mechanically extracted sample, which indicates more toughness and less flexibility of the fiber. 215 FTIR analysis In general, the major components of the plant fibers are cellulose, hemicellulose, and lignin. These components are mainly composed of alkanes, aromatics, alcohols, esters, and ketones with oxygencontaining functional groups. Hence, the FTIR analysis was performed on the extracted banana fibers to study the nature of functional groups present in the fibers. FTIR spectra depicted in 220 Figure 3 reflect the effectiveness of different retting processes. All the fiber bundles obtained by retting methods at several exposure times exhibit similar vibrations to those observed in fiber bundles obtained by mechanical extraction. Therefore, both retted and mechanically extracted fiber bundles present a similar chemical composition. Similar observations have been reported by Ganan et al. (2004) during the evaluation of chemical composition by FTIR spectroscopy for banana 225 fibers obtained using biological retting and mechanical extraction. An absorption band 3777 cm−1 corresponds to the stretching vibrations of O–H hydroxyl bonds. The broad absorption band at 3425 cm−1 corresponds to O–H stretching vibration of cellulose. The absorbance at 2928 cm−1 related to the C–H stretching vibrations of cellulose. The narrow band at 2357 cm−1 is associated with the presence of (NH2+ and NH+) amine groups. The band at 1594 cm−1 corresponds to C–C 230 stretching aromatic skeletal vibrations of lignin groups. The narrow band at 1404 cm−1 results in the presence of NH3+ amine salts. The absorbance at 1211 cm−1 is attributed to the C–O–C stretching vibrations of cellulose, hemicellulose, and lignin, suggesting that the fiber sample contains ethereal

Figure 3. FTIR spectra of BPF extracted by different methods.

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Table 3. FTIR spectra of BPF extracted by different methods. Wave number (cm−1) 3777 3425 2928 2357 1594 1404 1121 669 (w)

(s) (s) (w) (s) (vw) (m) (s)

Assignmenta O–H stretching O–H stretching C–H stretching NH2+, NH+ C=C aromatic skeletal vibrations NH3+ amine salt C–O–C asymmetric stretching C–S stretching

Origin Water, cellulose, hemicellulose, lignin Water, cellulose, hemicellulose, lignin Cellulose, hemicellulose, lignin Unknown compound Lignin Unknown compound Cellulose, hemicellulose, lignin Thiol, sulfide

Band intensity: s, strong; w, weak; m, medium; vw, very weak Pereira et al. (2014); Saba et al. (2015); Memon et al. (2008); Xu et al. (2015).

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character. The absorbance at 669 cm−1 is associated with the presence of C–S thiol and sulfide groups. The spectral analysis data of BPF samples by different methods are given in Table 3. 235 SEM and EDX analysis In view of the tex value of chemical retted fiber and its suitability in textile industries, chemical retted BPF alone was analyzed using FE-SEM with EDX analysis, and it was compared with mechanically extracted BPF of Xu et al. (2015). The SEM image (Figure 4a–4c) shows the longitudinal view of fiber bundles under different magnification. Further, it was observed that fiber 240 obtained from chemical retting process illustrates cleaner, rougher, and fibrillated fiber surface topography, which is due to the removal of hemicellulose, pectin, and lignin content, whereas mechanically extracted fiber had a smooth surface. The diameter of the chemical retted fiber was found to be approximately 180 µm. Alkali treatment reduces fiber diameter and helps in developing

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Figure 4. SEM analysis of BPF obtained from chemical retting method under different magnifications: (a) 959×; (b) 1.2K×; (c) 2.5K×; (d) EDX spectrum of BPF obtained from chemical retting.

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rough topography (Bessadok et al. 2009). The void spaces in the alignment of cellulose layers with 245 rough surface were also observed in the fiber bundles due to the removal of noncellulosic content such as pectin and lignin. Xu et al. (2015) reported that mechanically extracted fibers consist of waxy substances, hemicelluloses, cellulose, and lignin. A small number of surface cracks could also be observed in mechanically extracted fiber, which was absent in chemical retted fiber sample. EDX spectrum shows higher amount of oxygen, which indicates the presence of cellulose, impurities such 250 as Na and P present in least amount due to the chemical agents added during the retting process (Figure 4d).

Conclusion

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Extraction processes of banana fibers were performed by different methods that include mechanical extraction, chemical, microbial, and enzymatic retting. Each method has its advantages and disadvantages in the contexts of the yield, quality, and properties of fiber bundles obtained. Mechanical and chemical analysis of fibers helped to determine its applicability in various industries. FTIR analysis confirmed no major difference in the functional groups of banana fibers but intensity of the peak varies. The rough surface topography and increased aspect ratio due to the reduced fiber diameter proffer improved mechanical properties and fiber–matrix interface adhesion of chemical retted BPF. Selection of the appropriate extraction method is most important if the fibers are employed for textiles in making garments. The tensile, flexural properties, and the SEM analysis confirmed that chemical retting process using sodium hydroxide yielded fiber samples of low tex value (6.4), which would be more suitable for textile industries. Microbially retted fibers could be better employed for fiber reinforcement composites due to their higher lignin content. During the retting process, excessive degradation of the fiber material should be avoided, which will affect the fiber strength. Hence, the correlation between retting time and fiber quality should be investigated in future studies. Optimization of parameters and process upscaling is expected in future research to promote investment and greater use of banana fiber in various industries, thereby opening up new market niches for banana fiber.

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Acknowledgments The authors are thankful to the director, ICAR-NRC for Banana, Trichy, for granting the permission to do the project work at ICAR- NRCB and providing the necessary facilities to carry out this investigation. They also acknowledge the assistance provided by Shri. K. Kamaraju, Technical Assistant and Ms. A. Lilly Hilda for carrying out this piece of work. 275

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